Insulin plays an essential role in the regulation of the intermediary metabolism of carbohydrates, proteins, and fats in the liver, adipose tissue, and muscle. Insulin is only synthesized in pancreatic islet beta cells. The results from studies conducted in our laboratory and elsewhere strongly suggest that the PDX-1 homeoprotein is a key transcriptional activator of the insulin gene. This factor also appears to be required for transcription of genes important in pancreatic differentiation. Thus, the pancreas does not develop in the absence of PDX-1 expression. We propose to continue our ongoing studies aimed at determining how PDX-1 stimulates insulin gene transcription. Activation by PDX-1 appears to be mediated through cooperative interactions with the E2A-encoded basic helix-loop- helix proteins (E12, E47, or E2/5) that compose a part of the ICE activator, another essential insulin gene transcription factor. These studies will concentrate on identifying and characterizing the sequences within the PDX-1 protein important in this response. In addition, we will characterize the regulatory domain in PDX-1 protein important in this response. In addition, we will characterize the regulatory domain in PDX-1 that influences its transactivation potential in beta cells. Lastly, we will attempt to identify the factors that are necessary for controlling islet beta cell-specific transcription of the PDX-1 gene.